Loss of Dnmt1 catalytic activity reveals multiple roles for DNA methylation during pancreas development and regeneration

Department of Biochemistry and Biophysics, Programs in Developmental Biology, Genetics, and Human Genetics, Diabetes Center, and Liver Center, University of California, San Francisco, San Francisco, CA 94158-2324, USA. (
Developmental Biology (Impact Factor: 3.55). 08/2009; 334(1):213-23. DOI: 10.1016/j.ydbio.2009.07.017
Source: PubMed


Developmental mechanisms regulating gene expression and the stable acquisition of cell fate direct cytodifferentiation during organogenesis. Moreover, it is likely that such mechanisms could be exploited to repair or regenerate damaged organs. DNA methyltransferases (Dnmts) are enzymes critical for epigenetic regulation, and are used in concert with histone methylation and acetylation to regulate gene expression and maintain genomic integrity and chromosome structure. We carried out two forward genetic screens for regulators of endodermal organ development. In the first, we screened for altered morphology of developing digestive organs, while in the second we screed for the lack of terminally differentiated cell types in the pancreas and liver. From these screens, we identified two mutant alleles of zebrafish dnmt1. Both lesions are predicted to eliminate dnmt1 function; one is a missense mutation in the catalytic domain and the other is a nonsense mutation that eliminates the catalytic domain. In zebrafish dnmt1 mutants, the pancreas and liver form normally, but begin to degenerate after 84 h post fertilization (hpf). Acinar cells are nearly abolished through apoptosis by 100 hpf, though neither DNA replication, nor entry into mitosis is halted in the absence of detectable Dnmt1. However, endocrine cells and ducts are largely spared. Surprisingly, dnmt1 mutants and dnmt1 morpholino-injected larvae show increased capacity for pancreatic beta cell regeneration in an inducible model of pancreatic beta cell ablation. Thus, our data suggest that Dnmt1 is dispensable for pancreatic duct or endocrine cell formation, but not for acinar cell survival. In addition, Dnmt1 may influence the differentiation of pancreatic beta cell progenitors or the reprogramming of cells toward the pancreatic beta cell fate.

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Article: Loss of Dnmt1 catalytic activity reveals multiple roles for DNA methylation during pancreas development and regeneration

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    • "s a key role in allowing DNA methylation during the maturation of germ cells (Tang et al., 2009). MBPs belong to three different families of proteins: namely the MBD family (proteins containing a methyl- CpG - binding domain or MBD ) , the Kaiso and Kaiso - like proteins , and the SRA domain proteins ( Fournier et al., 2011 ) . In et al . , 2007 ; Anderson et al . , 2009 ; Smith et al . , 2011 ) . For instance , the dynamic changes in methylation seen during the course of mammalian embryonic development have been demonstrated in zebrafish embryos ( Mackay et al. , 2007 ) . Until now eight different dnmt genes are reported in zebrafish . Like mammals they have only one dnmt1 , and one dnmt2 , while other"
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    ABSTRACT: As a sequel of our investigations on the impact of epigenome in inducing fetal alcohol spectrum disorder (FASD) phenotypes in Japanese rice fish, we have investigated on several DNA methylation machinery genes including DNA methyl transferase 3ba (dnmt3ba) and methyl binding proteins (MBPs), namely, mbd1b, mbd3a, mbd3b, and mecp2 at transcription level. Studies were made during normal development, from 0 day post fertilization (dpf) to hatching, and also exposing the fertilized eggs to ethanol or a DNMT inhibitor, 5-azacytidine (5-azaC). We observed that during development, all these genes followed distinct expression patterns, generally high mRNA copies in early phases (0-1 dpf) and significantly low mRNA copies prior to or after hatching. Ethanol (100-500 mM, 0-2 dpf) was unable to alter any of these mRNAs in 2 dpf; additional four days (2-6 dpf) maintenance of these embryos in ethanol-free environment, on 6 dpf, were also unable to establish any significant difference in these mRNA levels in comparison with the corresponding controls. However, continuous exposure of fertilized eggs in 300 mM ethanol, 0-6 dpf, showed significantly high mRNA copies only in MBPs (mbd1b, mbd3a, mbd3b, mecp2). 5-azaC (2 mM) on 2 dpf was able to enhance only mbd3b mRNA. Removal of 5-azaC and maintenance of these embryos in clean medium, 2-6 dpf, showed significantly enhanced mbd3b and mecp2 mRNAs compared to corresponding controls on 6 dpf. Our studies showed that in Japanese rice fish embryogenesis both ethanol and 5-azaC has the potential to specifically modulate the developmental rhythm of DNA methylation machineries.
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    • "Embryos were sorted for GFP expression within 4 h of heat shock; only the brightest embryos were assessed for liver size on 5 dpf. Dnmt1 s904 mutants (Anderson et al., 2009) were obtained from M. Goll (Memorial Sloan-Kettering Cancer Center, New York, USA). Tg(fabp10:dsRed) fish (Dong et al., 2007) were obtained from D. Stainier. "

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    • "A more global analysis of zygotic transcription initiation in embryos in which maternal Dnmt1 has been abolished would lend support to this factor being the elusive transcriptional repressor. Early embryonic development is normal in zebrafish dnmt1 mutants, suggesting that zygotic dnmt1 has little if any role in controlling the MZT (Anderson et al., 2009; Goll et al., 2009). However, the MZT has not been studied in zebrafish embryos in which both maternal and zygotic dnmt1 function has been disrupted. "
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    ABSTRACT: The initial phases of embryonic development occur in the absence of de novo transcription and are instead controlled by maternally inherited mRNAs and proteins. During this initial period, cell cycles are synchronous and lack gap phases. Following this period of transcriptional silence, zygotic transcription begins, the maternal influence on development starts to decrease, and dramatic changes to the cell cycle take place. Here, we discuss recent work that is shedding light on the maternal to zygotic transition and the interrelated but distinct mechanisms regulating the onset of zygotic transcription and changes to the cell cycle during early embryonic development.
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